The CZECHII mouse colony represents a unique experimental model for identifying insertional mutations caused by mouse mammary tumor virus (MMTV) because they lack endogenous germline MMTV proviral sequences which are present in all other laboratory strains of mice. CZECHII mice and their MMTV-induced hyperplasias are utilized to identify early- acting genes and, subsequently, to determine which MMTV-induced mutations might be involved with progression from hyperplasia to tumor to metastasis. By taking advantage of the clonal dominant nature of the preneoplasias, their tumors and metastases, we hope to identify genes commonly affected by MMTV during these mutagenic events., and differentiation of normal mammary tissue in vivo.Using this experimental model, a new transforming INT gene (INT6) was identified in a premalignant CZECH mouse mammary outgrowth.This gene has recently been linked to the eIF3 translation initiation complex, a large multiple- subunit protein that plays a central role in the pathway of initiation by promoting binding of t-RNA and mRNA to the 40S ribosomal subunit. The INT6 gene has been relatively unchanged throughout evolution from flies to humans. Two growth factors, TGF-a and TGF-b1, were shown to exert opposing effects on mammary epithelial growth, survival and tumorigenesis in transgenic mouse models in collaborative studies with Glenn Merlino (LMB, NCI). In my studies on the identification, characterization and analyses of mammary epithelium-specific stem cells, several important breakthroughs have been made. We have demonstrated the existence of clonogenic progenitor cells among the mammary epithelium capable of producing lobular growth but not ductal branching morphogenesis and, conversely, progenitors capable of producing ductal morphogenesis but not lobulo- genesis. These cells are present in small numbers among the mammary population and appear to be produced by a third clonogenic cell which is capable of producing an entire functional mammary gland and replicating itself in the process. With Dr. G. Chepko (LTIB), the in situ morphological candidates for these three morphogenetically distinct mammary progenitors have been described by electron microscopy and their position within the tissue and change in number during mammary functional differentiation and involution specified. Preliminary studies demonstrate that a single mammary tissue- specific stem cell can regenerate an entire functional gland in vivo including the three types of clonogenic epithelial cells described above.These multipotent, proliferation-competent epithelial cells represent the immediate targets for oncogenic transformation in the mammary gland.